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1.
Clin J Pain ; 36(4): 296-301, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31977369

RESUMO

OBJECTIVES: The objectives of this study were to investigate the correlations between the minimum effective volume (MEV) of lidocaine 1.5% for an ultrasound-guided popliteal sciatic nerve block and individual factors including the cross-sectional nerve area, sex, age, body mass index, and the depth of the sciatic nerve and to evaluate the safety of combined femoral and sciatic nerve blocks by monitoring the plasma concentration of local anesthetics. METHODS: Forty patients received combined single-shot femoral and continuous sciatic nerve blocks. The femoral nerve block was performed with an in-plane technique and 15 mL of lidocaine 1.5%. A continuous peripheral nerve block annular tube was positioned between the tibial and peroneal nerves inside the paraneural sheath. Thirty minutes after the femoral nerve block, a loading dose of 5 mL of lidocaine 1.5% was given to block the sciatic nerve after obtaining the maximum compound muscle action potential (CMAP) amplitude using nerve conduction studies. Additional lidocaine 1.5% was pumped at a rate of 30 mL/h through the indwelling annular tube if, after 8 minutes, the CMAP amplitude was still present. The CMAP amplitude monitored by the nerve conduction studies and pinprick tests were recorded every 2 minutes after the administration of lidocaine 1.5%. When the CMAP amplitude decreased to nearly 0 mV, this MEV was recorded. The influences of the cross-sectional area of the sciatic nerve, sex, age, body mass index, and the depth of the sciatic nerve on the MEV were analyzed using stepwise multiple linear regression. Blood samples were collected from 10 patients to evaluate the safety of combined femoral and sciatic nerve blocks by ultra-performance liquid chromatography-tandem mass spectrometry. Blood was drawn at 0 minutes before femoral nerve injection; 0 minutes before sciatic nerve injection; 8 minutes after sciatic nerve injection; and 0, 10, 20, 30, 45, 60, 75, 90, and 120 minutes after the pumping of lidocaine 1.5% stopped. RESULTS: A significant correlation was found between the MEV of lidocaine 1.5% and the cross-sectional area of the sciatic nerve (r=0.459), with a regression equation of the MEV (mL)=5.969+0.095×(the cross-sectional area of the sciatic nerve). The coefficient of determination was 0.211 (P<0.05). The MEV of lidocaine 1.5% for complete sciatic nerve blocks ranged from 7 to 15 mL. The maximum concentrations of lidocaine, monoethylglycinexylidide, and glycinexylidide were 1672.9 (227.6), 265.7 (32.7), and 42.2 (22.4) ng/mL, respectively. CONCLUSIONS: There is a positive correlation between the cross-sectional area of the sciatic nerve and the MEV. The regression equation can help to predict the MEV of lidocaine 1.5% for popliteal sciatic nerve blocks. The maximum concentrations of lidocaine and its metabolites did not approach toxic threshold limits in this study.

2.
Nat Commun ; 10(1): 4285, 2019 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-31537803

RESUMO

The preparation of fluorescent discrete supramolecular coordination complexes (SCCs) has attracted considerable attention within the fields of supramolecular chemistry, materials science, and biological sciences. However, many challenges remain. For instance, fluorescence quenching often occurs due to the heavy-atom effect arising from the Pt(II)-based building block in Pt-based SCCs. Moreover, relatively few methods exist for tuning of the emission wavelength of discrete SCCs. Thus, it is still challenging to construct discrete SCCs with high fluorescence quantum yields and tunable fluorescence wavelengths. Here we report nine organoplatinum fluorescent metallacycles that exhibit high fluorescence quantum yields and tunable fluorescence wavelengths through simple regulation of their photoinduced electron transfer (PET) and intramolecular charge transfer (ICT) properties. Moreover, 3D fluorescent films and fluorescent inks for inkjet printing were fabricated using these metallacycles. This work provides a strategy to solve the fluorescence quenching problem arising from the heavy-atom effect of Pt(II), and offers an alternative approach to tune the emission wavelengths of discrete SCCs in the same solvent.

3.
BMC Complement Altern Med ; 19(1): 193, 2019 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-31362740

RESUMO

BACKGROUND: Wenshen Jianpi recipe (WSJPR), a blended traditional Chinese medicine, is considered to have the possible beneficial effect on the progression of diabetic nephropathy (DN). This present study was designed to elucidate this protective activity in a rat model with streptozotocin (STZ)-induced DN and to explore the possible underlying mechanism. METHODS: Adult Sprague Dawley (SD) rats were induced to develop DN through intraperitoneal injection of STZ (60 mg/kg). Animals were orally administered saline, WSJPR at 7.5, 15, 30 g/kg, and valsartan (25 mg/kg) daily for 8 weeks. Blood and 24-h urine samples of each rat were collected for biochemical examination at 2-week intervals. Microcirculatory blood flow in the renal cortex and hemorheology index were also measured. At the end of 8 weeks, all rats were sacrificed to obtain the kidney tissues for histological examination and reverse transcription polymerase chain reaction (RT-PCR) was used to analyze the transcriptional levels of nephrin and podocin genes. RESULTS: WSJPR could improve serum total protein (TP) and albumin (ALB), reduce the excretion rates of urine-TP (U-TP), urine-ALB (U-ALB) and urine urea nitrogen (UUN) (P < 0.05), although it did not significantly alter the hyperglycemia. In addition, treatment with WSJPR could strongly reduce blood flow, erythrocyte aggregation index, and ameliorate microcirculation. In histological measurement, WSJPR-treated rats showed a significant amelioration in glomerular hypertrophy and mesangial expansion. By RT-PCR, we found WSJPR up-regulated the nephrin and podocin expression at mRNA levels. CONCLUSION: This study suggested that WSJPR could effectively relieve renal damage and improve renal function of DN rats by ameliorating metabolism disorder and increasing the gene expression of nephrin and podocin, which might be a useful approach for the treatment of DN.


Assuntos
Nefropatias Diabéticas/tratamento farmacológico , Medicamentos de Ervas Chinesas/administração & dosagem , Proteinúria/tratamento farmacológico , Animais , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas/química , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Rim/efeitos dos fármacos , Rim/lesões , Rim/metabolismo , Masculino , Medicina Tradicional Chinesa , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteinúria/genética , Proteinúria/metabolismo , Ratos , Ratos Sprague-Dawley
4.
Phys Chem Chem Phys ; 21(5): 2365-2371, 2019 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-30666332

RESUMO

Fluorescent proteins (FPs) have become fundamental tools for live cell imaging. Most FPs currently used are members of the green fluorescent protein super-family, but new fluorophores such as bilin-FPs are being developed and optimized. In particular, the UnaG FP incorporates bilirubin (BR) as a chromophore, enhancing its fluorescence quantum yield by three orders of magnitude relative to that in solution. To investigate the mechanism of this dramatic enhancement and provide a basis for further engineering of UnaG and other tetrapyrrole-based fluorophores, we performed picosecond fluorescence and femtosecond transient absorption measurements of BR bound to UnaG and its N57A site-directed mutant. The dynamics of wt-UnaG, which has a fluorescence QY of 0.51, are largely homogeneous, showing an excited state relaxation of ∼200 ps, and a 2.2 ns excited-state lifetime decay with a kinetic isotope effect (KIE) of 1.1 for D2O vs. H2O buffer. In contrast, for UnaG N57A (fluorescence QY 0.01) the results show a large spectral inhomogeneity with excited state decay timescales of 47 and 200 ps and a KIE of 1.4. The non-radiative deactivation of the excited state is limited by proton transfer. The loss of direct hydrogen bonds to the endo-vinyl dipyrrinone moiety of BR leads to high flexibility and structural heterogeneity of UnaG N57A, as seen in the X-ray crystal structure.


Assuntos
Bilirrubina/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Bilirrubina/química , Bilirrubina/efeitos da radiação , Sítios de Ligação , Fluorescência , Proteínas de Fluorescência Verde/química , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/efeitos da radiação , Ligações de Hidrogênio , Luz , Mutação , Ligação Proteica
5.
Int J Mol Sci ; 19(9)2018 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-30201867

RESUMO

Current treatment of rheumatoid arthritis (RA) is limited by relative shortage of treatment targets. HM-3 is a novel anti-RA polypeptide consisting of 18 amino acids with integrin αVß3 and α5ß1 as targets. Previous studies confirmed that HM-3 effectively inhibited the synovial angiogenesis and the inflammatory response. However, due to its short half-life, the anti-RA activity was achieved by frequent administration. To extend the half-life of HM-3, we designed a fusion protein with name HM-3-Fc, by combination of modified Fc segment of immunoglobulin 4 (IgG4) with HM-3 polypeptide. In vitro cell experiments demonstrated that HM-3-Fc inhibited the proliferation of splenic lymphocytes and reduced the release of TNF-α from macrophages. The pharmacodynamics studies on mice paw in Collagen-Induced Arthritis (CIA) model demonstrated that HM-3-Fc administered once in 5 days in the 50 and 25 mg/kg groups, or once in 7 days in the 25 mg/kg group showed a better protective effect within two weeks than the positive control adalimumab and HM-3 group. Preliminary pharmacokinetic studies in cynomolgus confirmed that the in vivo half-life of HM-3-Fc was 15.24 h in comparison with 1.32 min that of HM-3, which demonstrated that an Fc fusion can effectively increase the half-life of HM-3 and make it possible for further reduction of subcutaneous injection frequency. Fc-HM-3 is a long-acting active molecule for RA treatment.


Assuntos
Artrite Experimental/prevenção & controle , Integrinas/antagonistas & inibidores , Linfócitos/citologia , Proteínas Recombinantes de Fusão/administração & dosagem , Baço/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Adalimumab/administração & dosagem , Adalimumab/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Meia-Vida , Humanos , Fragmentos Fc das Imunoglobulinas/administração & dosagem , Fragmentos Fc das Imunoglobulinas/farmacologia , Integrina alfa5beta1/antagonistas & inibidores , Integrina alfaVbeta3/antagonistas & inibidores , Linfócitos/efeitos dos fármacos , Camundongos , Peptídeos/administração & dosagem , Peptídeos/farmacologia , Proteínas Recombinantes de Fusão/farmacologia , Peixe-Zebra
6.
Acta Trop ; 183: 14-18, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29605156

RESUMO

The sensitivity and specificity are two crucial aspects of addressing the efficacy of diagnostic antigens. Achilles' heel of low sensitivity rate exists in current diagnostic recombinant antigens for schistosomiasis detection. This study focused on the diagnosis of water buffalo schistosomiasis japonica and a perspective of improving recombinant antigens' sensitivity was assessed using archived 220 water buffalo sera (114 positive sera, 92 negative sera and 14 Paramphistomum-infected sera) and the method of enzyme-linked immunosorbent assay (ELISA). The subjects included two trivalent recombinant proteins, one bivalent antigen and two single-molecular antigens. The crude antigen SEA (soluble egg antigen) was employed as reference antigen. The highest sensitivity rate in the five recombinant antigens assigned to the trivalent multi-epitope antigen PA4 (95.61%, 109/114), no significant difference with SEA (100%, 114/114, p = .836), and showing remarkable differences with the two single-molecular antigens (p < 0.01). In term of specificity, two trivalent multi-epitope antigens PA4 (97.83%, 90/92), PA5 (100%, 92/92) and the bivalent antigen PA3 (98.91%, 91/92) had few differences with one monovalent antigens PA1 (97.83%, 90/92, p = .304/0.103/0.640), significant differences with another monovalent antigens PA2 (92.39%, 85/92, p < 0.01) and SEA (82.61%, 76/92, p < 0.01). Additional, all the recombinant antigens had low cross-reactivity (7.14%, 1/14, 0% for PA5) with serum samples of paramphistomiasis, contrast with that of SEA (50%, 7/14, p < 0.01). The results indicated that multi-epitope antigens have the possibility to improve diagnostic sensitivity and the trivalent multi-epitope antigen PA4 possesses greater likelihood to be a diagnostic antigen for water buffalo schistosomiasis.


Assuntos
Anticorpos Anti-Helmínticos/sangue , Antígenos de Helmintos/sangue , Proteínas Recombinantes/sangue , Esquistossomose Japônica/sangue , Animais , Búfalos , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática/métodos , Epitopos/imunologia , Proteínas Recombinantes/imunologia , Esquistossomose Japônica/imunologia , Sensibilidade e Especificidade , Testes Sorológicos
7.
Reprod Biol Endocrinol ; 16(1): 16, 2018 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-29482568

RESUMO

Asthenozoospermia is considered as a common cause of male infertility and characterized by reduced sperm motility. However, the molecular mechanism that impairs sperm motility remains unknown in most cases. In the present review, we briefly reviewed the proteome of spermatozoa and seminal plasma in asthenozoospermia and considered post-translational modifications in spermatozoa of asthenozoospermia. The reduction of sperm motility in asthenozoospermic patients had been attributed to factors, for instance, energy metabolism dysfunction or structural defects in the sperm-tail protein components and the differential proteins potentially involved in sperm motility such as COX6B, ODF, TUBB2B were described. Comparative proteomic analysis open a window to discover the potential pathogenic mechanisms of asthenozoospermia and the biomarkers with clinical significance.


Assuntos
Astenozoospermia/metabolismo , Proteoma/metabolismo , Espermatozoides/metabolismo , Humanos , Masculino , Proteômica , Motilidade Espermática/fisiologia
8.
Anal Chem ; 90(3): 2018-2022, 2018 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-29275628

RESUMO

Raman scattering and fluorescence spectroscopy permeate analytic science and are featured in the plasmon-enhanced spectroscopy (PES) family. However, the modest enhancement of plasmon-enhanced fluorescence (PEF) significantly limits the sensitivity in surface analysis and material characterization. Herein, we report a Ag nanoantenna platform, which simultaneously fulfills very strong emission (an optimum average enhancement of 105-fold) and an ultrafast emission rate (∼280-fold) in PES. For applications in surface science, this platform has been examined with a diverse array of fluorophores. Meanwhile, we utilized a finite-element method (FEM) and time-dependent density functional theory (TD-DFT) to comprehensively investigate the mechanism of largely enhanced radiative decay. PES with a shell-isolated Ag nanoantenna will open a wealth of advanced scenarios for ultrasensitive surface analysis.

9.
Oncotarget ; 8(33): 55646-55656, 2017 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-28903451

RESUMO

Sperm morphology displays a potential impact on sperm function and may ultimately impact reproductive function. Current studies have investigated the correlation between sperm morphology with unexplained recurrent spontaneous abortion (RSA) but have shown inconsistent results. Hence, we systematically searched MEDLINE, EMBASE, CNKI databases, as well as the Cochrane Library for studies that examined the association between sperm morphology and unexplained RSA. Fifteen studies were identified, including 883 cases and 530 controls. Our meta-analysis results indicated that the percentage of normal sperm morphology from men with RSA partners was significantly lower than those from normal controls(SMD [95% CI]: - 0.60 [-0.81, -0.40]; P<0.00001) and the percentage of sperm morphologic alterations was significantly higher in patients with RSA compared with the control group (SMD [95% CI]: 0.92 [0.42, 1.43]; P=0.0004). The present study suggested that the percentage of normal sperm morphology may indeed decrease in men from RSA group compared with controls. However, there were some limitations in the study such as the differences in stain techniques and classification criteria. Further evidences are needed to better elucidate the relationship between sperm morphology and unexplained RSA.

10.
Anal Bioanal Chem ; 409(21): 5073-5080, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28687887

RESUMO

pH-dependent protein adsorption on mesoporous silica nanoparticle (MSN) was examined as a unique means for pH monitoring. Assuming that the degree of protein adsorption determines the distance separating protein molecules, we examined the feasibility of nanoscale pH probes based on fluorescence resonance energy transfer (FRET) between two fluorescent proteins (mTurquoise2 and mNeonGreen, as donor and acceptor, respectively). Since protein adsorption on MSN is pH-sensitive, both fluorescent proteins were modified to make their isoelectric points (pIs) identical, thus achieving comparable adsorption between the proteins and enhancing FRET signals. The adsorption behaviors of such modified fluorescent proteins were examined along with ratiometric FRET signal generation. Results demonstrated that the pH probes could be manipulated to show feasible sensitivity and selectivity for pH changes in hosting solutions, with a good linearity observed in the pH range of 5.5-8.0. In a demonstration test, the pH probes were successfully applied to monitor progress of enzymatic reactions. Such an "in situ-assembling" pH sensor demonstrates a promising strategy in developing nanoscale fluorescent protein probes. Graphical abstract Working principle of the developed pH sensor TNS; and FRET Ratio (I528/I460) as a function of pH under different protein feed ratios (mNeonGreen to mTurquoise2).


Assuntos
Transferência Ressonante de Energia de Fluorescência/métodos , Corantes Fluorescentes/química , Proteínas Luminescentes/química , Nanopartículas/química , Adsorção , Estudos de Viabilidade , Concentração de Íons de Hidrogênio
11.
Sci Rep ; 7(1): 5212, 2017 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-28701752

RESUMO

Schistosomiasis caused by schsitosomes is a serious global public health concern. The tegument that surrounds the worm is critical to the schistosomes survival. The tegument apical membrane undergoes a continuous process of rupture and repair owing to membranous vacuoles fusing with the plasma membrane. Vesicle-associated membrane protein 2 (VAMP2), a member of soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNAREs) is required for membrane fusion. Here, we used RNA interference (RNAi) to knock down the expression of VAMP2 of Schistosoma japonicum (SjVAMP2), and both real-time PCR and western blot analysis confirmed the suppression of this molecule, as well as the suppression of the transcript levels of schistosome glucose transporters (SGTP1 and SGTP4), and insulin receptors (SjIR1 and SjIR2). SjVAMP2-suppressed worms exhibited a lower viability, and phenotypic alterations were also observed in the tegument. Moreover, the glucose consumption of SjVAMP2-suppressed worms decreased significantly in 4 and 6 days, respectively, as well as a significant reduction in egg production. We also observed a significant reduction in worm burden and hepatic eggs burden in two independent RNAi experiment in vivo, and minor pathological changes in mice treated with SjVAMP2 specific small interfering (si)RNA. These findings reveal that SjVAMP2 may play important roles in the maintenance of tegument, glucose uptake, worm development and egg production in schistosomes.


Assuntos
Glucose/metabolismo , Reprodução , Schistosoma japonicum/anatomia & histologia , Schistosoma japonicum/crescimento & desenvolvimento , Esquistossomose Japônica/parasitologia , Proteína 2 Associada à Membrana da Vesícula/metabolismo , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Receptor de Insulina/genética , Receptor de Insulina/metabolismo , Schistosoma japonicum/metabolismo , Esquistossomose Japônica/metabolismo , Esquistossomose Japônica/patologia , Caramujos/parasitologia , Proteína 2 Associada à Membrana da Vesícula/genética
12.
Parasit Vectors ; 10(1): 288, 2017 Jun 09.
Artigo em Inglês | MEDLINE | ID: mdl-28599669

RESUMO

BACKGROUND: Schistosomiasis remains an important global public health problem, as millions of people are at risk of acquiring infection. An ideal method for sustainable control of schistosomiasis would be to develop an efficient vaccine. Schistosomes can survive in the host vascular system by immune evasion, regulating the host complement cascade. Schistosoma japonicum tetraspanning orphan receptor (SjTOR) is a complement regulator, which is a tegument membrane protein. To date there is no experimental evidence to explain the function of SjTOR. RESULTS: We cloned the first extracellular domain of the SjTOR (SjTOR-ed1) gene and expressed the gene in Escherichia coli. The expression level of SjTOR in different developmental stages of S. japonicum was assessed by quantitative real-time RT-PCR. Western blotting showed that recombinant SjTOR-ed1 (rSjTOR-ed1) could be recognised by schistosome-infected mouse serum. Immunolocalization indicated that the protein was mainly distributed on the tegument of the parasite. Haemolytic assays and ELISA revealed that rSjTOR-ed1 could inhibit complement hemolysis and bind to complement C2. Purified rSjTOR-ed1 emulsified with ISA206 adjuvant could induce a significant reduction of worm burden from 24.51 to 26.51%, and liver egg numbers from 32.92 to 39.62% in two independent trials in mice. CONCLUSIONS: The results of this study indicated that rSjTOR-ed1 could inhibit complement hemolysis and bind to complement C2, and it is a potential vaccine candidate that protects against S. japonicum infection.


Assuntos
Complemento C2/metabolismo , Proteínas de Helminto/metabolismo , Interações Hospedeiro-Patógeno , Fatores Imunológicos/metabolismo , Ribonucleases/metabolismo , Schistosoma japonicum/imunologia , Esquistossomose/imunologia , Animais , Modelos Animais de Doenças , Proteínas de Helminto/genética , Camundongos Endogâmicos C57BL , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Ribonucleases/genética , Células Th2/imunologia
13.
Sci Rep ; 7(1): 4209, 2017 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-28646144

RESUMO

In this paper, we propose a novel and sensitive ratiometric analysis method that uses the fractional intensities of time-resolved fluorescence of genetically encoded fluorescent NADH/NAD+ biosensors, Peredox, SoNar, and Frex. When the conformations of the biosensors change upon NADH/NAD+ binding, the fractional intensities (α i τ i ) have opposite changing trends. Their ratios could be exploited to quantify NADH/NAD+ levels with a larger dynamic range and higher resolution versus commonly used fluorescence intensity and lifetime methods. Moreover, only one excitation and one emission wavelength are required for this ratiometric measurement. This eliminates problems of traditional excitation-ratiometric and emission-ratiometric methods. This method could be used to simplify the design and achieve highly sensitive analyte quantification of genetically encoded fluorescent biosensors. Wide potential applications could be developed for imaging live cell metabolism based on this new method.


Assuntos
Técnicas Biossensoriais/métodos , NAD/análise , Fluorescência , Fatores de Tempo
14.
Infect Dis Poverty ; 6(1): 84, 2017 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-28388965

RESUMO

BACKGROUND: Schistosomiasis remains a major public health concern in China and an epidemiological survey has revealed that schistosome-infected bovines and goats are the main transmission sources for the disease. Therefore, development of a sensitive technique for the diagnosis of schistosomiasis in domestic animals is necessary. METHOD: A novel colloidal gold immunochromatography assay (GICA) strip was developed for detecting Schistosoma japonicum in domestic animals. The colloidal gold was conjugated with recombinant streptococcal protein G (rSPG). As the test and control lines, the schistosome soluble egg antigen and rSPG, respectively, were blotted on nitrocellulose membrane. RESULTS: The lowest detectable serum dilution was 1∶640 for schistosome-infected buffaloes. The cross-reaction rate of GICA was 14.29% with Paramphistomum sp. in buffaloes, 16.67% with Haemonchus sp. in goats, and 33.33% with Orientobilharzia sp. in goats. These results were slightly lower and similar to those obtained through ELISA. Moreover, the strips for detecting S. japonicum in mice, rabbits, buffaloes, and goats showed high sensitivity (100.00%, 100.00%, 100.00%, and 100.00%, respectively) and specificity (100.00%, 100.00%, 94.23%, and 88.64%, respectively). And the sensitivity or specificity of the GICA strips did not present any significant differences after storage for 12 months at room temperature. When compared with ELISA, the GICA strips exhibited similar sensitivity and specificity in the diagnosis of schistosomiasis in mice, rabbits, buffaloes, and goats. Besides, only 5 µl of serum are required for the test and the detection can be completed within 5 min. CONCLUSION: This study is the first to develop a GICA strip using gold-rSPG conjugate for the diagnosing of schistosomiasis in domestic animals, and preliminary results showed that the developed strip may be suitable for large-scale screening of schistosomiasis in endemic areas.


Assuntos
Anticorpos Anti-Helmínticos/sangue , Cromatografia de Afinidade/métodos , Coloide de Ouro/análise , Schistosoma japonicum/imunologia , Esquistossomose Japônica/veterinária , Animais , Animais Domésticos , China , Reações Cruzadas , Esquistossomose Japônica/diagnóstico , Sensibilidade e Especificidade
15.
Parasitol Res ; 116(4): 1361-1372, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28285327

RESUMO

Thioredoxin peroxidases (TPxs) play an important role in maintaining redox homeostasis and in protecting organisms from the accumulation of toxic reactive oxygen species (ROS). In this study, we isolated the thioredoxin peroxidase-3 gene of Schistosoma japonicum, SjTPx-3. The open reading frame (ORF) of SjTPx-3 was 663 bp encoding 220 amino acids with a molecular weight of 24.99 kDa and an isoelectric point of 6.20. Quantitative real-time reverse transcription-polymerase chain reaction indicated that SjTPx-3 was expressed in all different stages of the parasites, with highest expression in 35-day-old worms. The ORF of SjTPx-3 was subcloned into pET-32a (+) vectors and expressed in Escherichia coli. Recombinant SjTPx-3 (rSjTPx-3) was expressed as a soluble protein with good antigenicity, as demonstrated by western blotting. Immunohistochemical analysis revealed that SjTPx-3 was mainly localized on the tegument of the parasites. Mice vaccinated with rSjTPx-3 had a 37.02% (P < 0.05) reduction in worm burden and 56.52% (P < 0.05) reduction in liver egg production compared with control, unvaccinated mice. Enzyme-linked immunosorbent assay analysis demonstrated that rSjTPx-3 could induce high levels of anti-rSjTPx-3-specific IgG, IgG1, and IgG2a antibodies. Characteristic Th1 and Th2 immune response cytokines were detected by flow cytometry and were increased by rSjTPx-3. Taken together, these results suggest that SjTPx-3 is an antioxidant enzyme responsible for protecting S. japonicum from oxidative stress. rSjTPx-3 may represent a potential vaccine candidate and/or new drug target for patients with schistosomiasis.


Assuntos
Anticorpos Anti-Helmínticos/imunologia , Proteínas de Helminto/imunologia , Peroxirredoxinas/metabolismo , Schistosoma japonicum/metabolismo , Esquistossomose Japônica/prevenção & controle , Vacinas/imunologia , Animais , Clonagem Molecular , Feminino , Proteínas de Helminto/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Fases de Leitura Aberta , Peroxirredoxinas/imunologia , Esquistossomose Japônica/parasitologia
16.
Biotechnol Appl Biochem ; 64(4): 482-489, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27222443

RESUMO

This work examines the feasibility of using a pH-sensitive fluorescent protein as a molecular reporter for enzyme-catalyzed prodrug activation reaction. Specifically, a ratiometric pHluorins was examined for detection of the activity of horseradish peroxidase (HRP) for the activation of indole-3-acetic acid. The pHluorins and HRP were conjugated chemically, forming a biocatalyst with a self-reporting function. Results showed that the characteristic fluorescence intensity ratio of the conjugate shifted from 1.47 to 1.40 corresponding to the progress of the prodrug activation reaction. The effectiveness of applying the conjugate for inhibition of the growth of Bcap-37 cells was also demonstrated simultaneously with reaction monitoring. The results reveal a very promising approach to realizing in situ monitoring of enzyme activities based on pH shifting for enzyme-based prodrug therapy applications.


Assuntos
Biocatálise , Técnicas Biossensoriais , Proteínas de Fluorescência Verde/química , Peroxidase do Rábano Silvestre/metabolismo , Ácidos Indolacéticos/metabolismo , Sondas Moleculares/química , Pró-Fármacos/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ativação Enzimática , Proteínas de Fluorescência Verde/isolamento & purificação , Proteínas de Fluorescência Verde/metabolismo , Peroxidase do Rábano Silvestre/química , Humanos , Concentração de Íons de Hidrogênio , Ácidos Indolacéticos/química , Sondas Moleculares/metabolismo , Pró-Fármacos/química
17.
Zhonghua Nan Ke Xue ; 23(5): 431-435, 2017 May.
Artigo em Chinês | MEDLINE | ID: mdl-29717834

RESUMO

Objective: To investigate the relationship between hepatitis B virus (HBV) infection and the incidence of male immune infertility. METHODS: Based on the levels of serum HBsAg, 3 124 infertile men were classified into an HBV-positive and an HBV-negative group and, according to the results of IBT tests, those with immune infertility were further divided into an HBV-positive and an HBV-negative group. Statistical analyses were made on the incidence rate of immune infertility and seminal parameters in the immune infertility patients of the HBV-positive and HBV-negative groups, the correlation of the number of HBV DNA copies in the serum with that in the seminal plasma of the HBV-positive patients, the association of the numbers of HBV DNA copies in the serum and seminal plasma with semen parameters, and the relationship of the number of HBV DNA copies in the seminal plasma with the incidence of immune infertility. Sperm concentration and the percentage of progressively motile sperm (PMS) were measured by computer-aided sperm analysis, sperm morphology determined by Diff-Quik staining, the level of HBsAg detected by ELISA, and the numbers of HBV DNA copies in the serum and seminal plasma calculated by RT-PCR. RESULTS: The incidence rate of immune infertility was significantly higher in the HBV-positive than in the HBV-negative group (20.3 vs 3.3%, χ2 = 187.5, P <0.01), and the percentage of morphologically normal sperm (MNS) was markedly lower in the HBV-positive than in the HBV-negative infertility patients (ï¼»3.9 ± 1.7ï¼½ vs ï¼»6.3 ± 2.2ï¼½%, P <0.05), but no statistically significant differences were observed between the two groups of infertile males in the semen volume, sperm concentration, or PMS (P >0.05). The number of HBV DNA copies in the serum was positively correlated with that in the seminal plasma (rs = 0.86, P <0.01) while both the number of HBV DNA copies in the serum and that in the seminal plasma were negatively correlated with PMS (r = -0.233 and -0.465, P <0.01) and MNS (r = -0.250 and -0.508, P <0.01). The incidence rate of immune infertility showed no statistically significant differences among the groups with different numbers of HBV DNA copies in the seminal plasma (P >0.05). CONCLUSIONS: HBV infection can increase the incidence rate of immune infertility in men and is correlated with the low quality of sperm.


Assuntos
Hepatite B/complicações , Infertilidade Masculina/epidemiologia , Hepatite B/imunologia , Antígenos de Superfície da Hepatite B/análise , Vírus da Hepatite B/imunologia , Humanos , Incidência , Infertilidade Masculina/virologia , Masculino , Sêmen , Análise do Sêmen , Contagem de Espermatozoides
18.
Guang Pu Xue Yu Guang Pu Fen Xi ; 37(2): 476-80, 2017 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-30280537

RESUMO

Considering the important role of metal ions including copper ions are playing in human body, a novel single-Trp peptide WDAHSS was designed and synthesized in this study to achieve sensitive detection of copper ions via fluorescence spectroscopy. The intrinsic fluorescence of a tryptophan residue in WDAHSS, which was the only source of the molecular fluorescence, could be easily quenched with copper ions. By comparing fluorescence spectra of WDAHSS with those of tryptophan molecules at different pH values, the quenching mechanism of WDAHSS was explored in detail. Research showed that the histidine in WDAHSS bound copper ions with metal coordination. With participation of peptide bond, a square planar structure was formed. It was a consequent chelation of copper ions that caused the quenching of tryptophan residue. At the same time, this study discussed how pH conditions affected the fluorescence spectra of WDAHSS. Furthermore, association constants of copper ions towards WDAHSS were calculated through fluorescence measurements and fitting analyses. To enhance the anti-jamming ability to pH variation, the amino terminal of WDAHSS was intentionally acetylized, leading to a stable fluorescence emission under physiological pH conditions. Besides, WDAHSS was designed as a special structure to enhance the selectivity and biocompatibility of its sensitive detection of copper ions. Further studies on WDAHSS may help to improve the fluorescence imaging detection in vivo.


Assuntos
Fluorescência , Quelantes , Histidina , Íons , Metais , Peptídeos , Espectrometria de Fluorescência , Triptofano
19.
J Proteomics ; 148: 202-12, 2016 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-27535354

RESUMO

UNLABELLED: Lysine acetylation, a ubiquitous and conserved posttranslational modification, has recently been shown to participate in many diverse non-chromatin-associated biological processes in prokaryotes and eukaryotes. However, the full extent and functional significance of acetylation in Schistosoma japonicum is still unknown. To investigate the nature, extent, and biological functions of lysine acetylation in schistosomes, immunoaffinity-based acetyl-lysine peptide enrichment, integrated with mass spectrometry, was used to comprehensively characterize the lysine-acetylated proteins in this parasite. In total, 1109 acetylated proteins and 2393 acetylation sites in S. japonicum were identified, representing the largest acetylome yet reported in a parasite. In a bioinformatic analysis showed that these acetylated proteins were mainly enriched in the biological process categories of metabolism, gene expression, translation, and transport. The classification according to molecular function revealed that the largest class involved the catalytic activity of different enzymes, including oxidoreductase, transferase, and pyrophosphatase activities. Most of the acetylated proteins in the cellular component category occurred in the cytoplasm, membrane, cytoskeleton, and nucleus. These data demonstrate the generality of lysine acetylation and provide the first global survey of acetylation in schistosomes. Our findings are an exciting starting point for the further exploration of the functions of acetylation in the biology of this parasite. SIGNIFICANCE: Schistosomiasis is one of the world's most prevalent and neglected tropical parasitic zoonotic diseases, and it causes almost 200,000 deaths annually. To control and eradicate schistosomiasis, effective vaccines are urgently required, and drug targets that are essential for schistosome survival must be identified in fundamental studies of schistosome biology. Posttranslational modifications are complex, fundamental, and important mechanisms that regulate the physiological functions of organisms. Lysine acetylation, a ubiquitous and conserved posttranslational modification, has recently been shown to participate in many diverse non-chromatin-associated biological processes in prokaryotes and eukaryotes. However, the full extent and functional significance of acetylation in Schistosoma japonicum is still unknown. To investigate the nature, extent, and biological functions of lysine acetylation in S. japonicum, we employ immunoaffinity-based acetyl-lysine peptide enrichment, integrated with mass spectrometry to comprehensively characterize the lysine-acetylated proteins in this parasite. The results of our data demonstrate the generality of lysine acetylation and provide the first global survey of acetylation in schistosomes. Our findings are an exciting starting point for the further exploration of the functions of acetylation in the biology of this parasite. Meanwhile, identifying the mechanisms and proteins targeted by acetylation may also provide a promising avenue for specific drug design and the development of sophisticated therapeutic strategies.


Assuntos
Proteínas de Helminto/análise , Processamento de Proteína Pós-Traducional , Proteoma/metabolismo , Schistosoma japonicum/química , Acetilação , Animais , Transporte Biológico , Biologia Computacional , Expressão Gênica , Proteínas de Helminto/metabolismo , Lisina/metabolismo , Metabolismo , Biossíntese de Proteínas , Proteoma/análise , Espectrometria de Massas em Tandem
20.
Gene ; 592(1): 71-77, 2016 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-27461946

RESUMO

Schistosomiasis remains a serious public health concern in China. BALB/c mice are susceptible to Schistosoma japonicum infection, whereas the Wistar rats are less susceptible. Apoptosis phenomenon was observed in 42d adult worms of S. japonicum from both rats and mice at the morphologic, DNA, cellular, and gene levels by transmission electron microscopy (TEM), fluorometric terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) analysis, fluorescein isothiocyanate-annexin-V/propidium iodide staining flow cytometry (FCM) analysis, and real-time PCR. The results showed that the apoptotic state in worms from two different susceptible hosts was diverse. Several classical hallmarks of apoptosis, including cell shrinkage, chromatin condensation and lunate marginalization, splitting of the nucleoli, nuclear shrinkage and apoptotic body formation were observed by TEM. TUNEL analysis showed that there were much more apoptosis spots in adult worms from rats than those from mice. Statistical analysis revealed that the degree of apoptosis and percentage of necrotic cells in adult worms from Wistar rats were significantly greater (P<0.01) than those from BALB/c mice by flow cytometry. A total of 15 apoptosis-associated genes including the major components of an intrinsic cell-death pathway were identified from S. japonicum in this study, suggested that a similar apoptosis pathway might occur in S. japonicum. Real-time PCR analyses revealed that the expression levels of most of the tested apoptosis-associated genes, except CASP7, were significantly higher or at the similar level in adult worms from Wistar rats, as compared to those from BALB/c mice. The results obtained in this study collectively demonstrated that differential development of adult S. japonicum in less-susceptible rats and susceptible mice was significantly associated with apoptosis in the worm, and provided valuable information to guide further investigations of the mechanisms governing apoptosis and host interactions in schistosome infection.


Assuntos
Apoptose , Especificidade de Hospedeiro , Schistosoma japonicum/genética , Animais , Genes de Helmintos , Interações Hospedeiro-Parasita , Camundongos , Camundongos Endogâmicos BALB C , Ratos , Ratos Wistar , Schistosoma japonicum/isolamento & purificação , Schistosoma japonicum/metabolismo , Schistosoma japonicum/patogenicidade
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